Proteins are essential nutrients for consumers, especially those on plant-based diets. The use of macaúba pulp cake, a by-product of the macaúba oil industry, promotes sustainability and the valorization of agro-industrial waste, aligning with circular economy practices. This study aimed to investigate, develop, and optimize the production process of K. marxianus yeast extract from macaúba pulp cake for use as a protein ingredient in plant-based foods. K. marxianus yeast was chosen for its thermotolerance and ability to utilize different carbon sources, making it valuable for modern biotechnology. The yeast was cultivated in a medium containing dried macaúba pulp cake and subjected to two cell lysis processes: thermal shock and ultrasound. Characterization of the extract included analyses of total reducing sugars, total proteins, and microscopic observation. Cultivation of K. marxianus in a medium containing dried macaúba pulp cake was viable, resulting in significant cell growth. Analysis revealed that proteins in the cultivation medium mainly originated from the macaúba cake, highlighting the need to optimize cell lysis processes to efficiently release intracellular components of the yeast. Microscopic analyses showed no differences between control samples and post-thermal shock treatment, but some differences in cell density and integrity were observed between ultrasound treatments, suggesting potential success in cell disruption. This extract has great potential for the plant-based food industry, providing essential proteins and promoting sustainability by utilizing agro-industrial by-products.